Lab report on liver function test

1. KENYATTA UNIVERSITY
SCHOOL OF MEDICINE.
DEPARTMENT OF MEDICAL BIOCHEMESTRY
NAME: LANDO ELVIS OTIENO
REG NO: P29S/16344/2015.
COURSE: MBCHB
LECTURER: Prof Njogu
HANDING DATE: /3/2016
SUBJECT: PRACTICAL REPPORT ON TEST FOR DETERMINATION OF
LIVER FUNCTION TESTS.
3/1/2016
Lando Elvis
©

2. Abstract
Laboratory liver tests are broadly defined as tests useful in the evaluation and treatment of
patients with hepatic dysfunction. The liver carries out metabolism of carbohydrate, protein and
fats. Some of the enzymes and the end products of the metabolic pathway which are very
sensitive for the abnormality occurred may be considered as biochemical marker of liver
dysfunction. Some of the biochemical markers such as serum bilirubin, alanine amino
transferase, aspartate amino transferase, ratio of aminotransferases, alkaline phosphatase, gamma
glutamyl transferase, 5′ nucleotidase, ceruloplasmin, α-fetoprotein are considered in this article.
An isolated or conjugated alteration of biochemical markers of liver damage in patients can
challenge the clinicians during the diagnosis of disease related to liver directly or with some
other organs. The term “liver chemistry tests” is a frequently used but poorly defined phrase that
encompasses the numerous serum chemistries that can be assayed to assess hepatic function
and/or injury.
Alanine amino transferase (ALT)
ALT is found in kidney, heart, muscle and greater concentration in liver compared with other
tissues of the body. ALT is purely cytoplasmic catalysing the transamination reaction . Normal
serum ALT is 7–56 U/ L . Any type of liver cell injury can reasonably increases ALT levels.
Elevated values up to 300 U/L are considered nonspecific. Marked elevations of ALT levels
greater than 500 U/L observed most often in persons with diseases that affect primarily
hepatocytes such as viral hepatitis, ischemic liver injury (shock liver) and toxin-induced liver
damage. Despite the association between greatly elevated ALT levels and its specificity to
hepatocellular diseases, the absolute peak of the ALT elevation does not correlate with the extent
of liver cell damage . Viral hepatitis like A, B, C, D and E may be responsible for a marked
increase in aminotransferase levels. The increase in ALT associated with hepatitis C infection
tends to be more than that associated with hepatitis A or B . Moreover in patients with acute
hepatitis C serum ALT is measured periodically for about 1 to 2 years . Persistence of elevated
ALT for more than six months after an occurrence of acute hepatitis is used in the diagnosis of
chronic hepatitis. Elevation in ALT levels are greater in persons with nonalcoholic
steatohepatitis than in those with uncomplicated hepatic steatosis . In a recent study the hepatic
fat accumulation in childhood obesity and nonalcoholic fatty liver disease causes serum ALT
elevation. Moreover increased ALT level was associated with reduced insulin sensitivity,
adiponectin and glucose tolerance as well as increased free fatty acids and triglycerides .
Presence of Bright liver and elevated plasma ALT level was independently associated with
increased risk of the metabolic syndrome in adults . ALT level is normally elevated during 2nd
trimester in asymptomatic normal pregnancy . In one of the study, serum ALT levels in
symptomatic pregnant patients such as in hyperemesis gravidarum was 103.5U/L, in pre-
eclampsia patients was 115U/L and in haemolysis with low platelet count patients showed
149U/L. However in the same study ALT rapidly drops more than 50% of the elevated values
within 3 days indicating the improvement during postpartum . One of the recent study has shown

3. that coffee and caffeine consumption reduces the risk of elevated serum ALT activity in
excessive alcohol consumption, viral hepatitis, iron overload, overweight, and impaired glucose
metabolism
Principle of ALT test.
In the reaction, ALT catalyzes the reversible transamination of L-alanine and α-ketoglutarate to
pyruvate and L-glutamate. The pyruvate is then reduced to lactate in the presence of lactate
dehydrogenase (LDH) with the concurrent oxidation of NADH to NAD. The system monitors
the rate of change in absorbance at 340 nm over a fixedtime interval. The rate of change in
absorbance is directly proportional to the ALT activity in the sample.
Normal ranges:
Reference ranges Female≤ 34 IU/L Male≤ 52 IU/L
Aspartate amino transferase (AST)
AST catalyse transamination reaction. AST exist two different isoenzyme forms which are
genetically distinct, the mitochondrial and cytoplasmic form. AST is found in highest
concentration in heart compared with other tissues of the body such as liver, skeletal muscle and
kidney . Normal serum AST is 0 to 35U/L. Elevated mitochondrial AST seen in extensive tissue
necrosis during myocardial infarction and also in chronic liver diseases like liver tissue
degeneration and necrosis . About 80% of AST activity of the liver is contributed by the
mitochondrial isoenzyme, whereas most of the circulating AST activity in normal people is
derived from the cytosolic isoenzyme. However the ratio of mitochondrial AST to total AST
activity has diagnostic importance in identifying the liver cell necrotic type condition and
alcoholic hepatitis . AST elevations often predominate in patients with cirrhosis and even in liver
diseases that typically have an increased ALT . AST levels in symptomatic pregnant patient in
hyperemesis gravidarum were 73U/L, in pre-eclampsia 66U/L, and 81U/L was observed in
hemolysis with low platelet count and elevated liver enzymes.
Principle for AST test
In the reaction, the AST catalyzes the reversible transamination of L-aspartate and α-
ketoglutarate to oxaloacetate and L-glutamate. The oxaloacetate is then reduced to malate in the
presence of malate dehydrogenase with the concurrent oxidation of NADH to NAD. The system

4. monitors the rate of change in absorbance at 340 nm over a fixed-time interval. The rate of
change in absorbance is directly proportional to the AST activity in the sample.
Reference Values
Males
0-11 months: not established
1-13 years: 8-60 U/L
> or =14 years: 8-48 U/L
Females
0-11 months: not established
1-13 years: 8-50 U/L
> or =14 years: 8-43 U/L
AST/ALT ratio
The ratio of AST to ALT has more clinical utility than assessing individual elevated levels. A
coenzyme pyridoxal-5'-phosphate deficiency may depress serum ALT activity and consequently
increases the AST/ALT ratio . An elevated ratio greater than 1 shows advanced liver fibrosis and
chronic hepatitis C infection. However, an AST/ALT ratio greater than 2 characteristically is
present in alcoholic hepatitis. A recent study differentiated nonalcoholic steatohepatitis (NASH)
from alcoholic liver disease showing AST/ALT ratio of 0.9 in NASH and 2.6 in patients with
alcoholic liver disease. A mean ratio of 1.4 was found in patients with cirrhosis related to NASH.
Wilson's disease can cause the ratio to exceed 4.5 and similar such altered ratio is found even in
Hyperthyroidism .
Gamma Glutamyl Transferase (GGT)
GGT is a microsomal enzyme present in hepatocytes and biliary epithelial cells, renal tubules,
pancreas and intestine. It is also present in cell membrane performing transport of peptides into
the cell across the cell membrane and involved in glutathione metabolism. Serum GGT activity
mainly attributed to hepatobiliary system even though it is found in more concentration in renal
tissue . The normal level of GGT is 9 to 85 U/L. In acute viral hepatitis the levels of GGT will
reach the peak in the second or third week of illness and in some patients remain elevated for 6
weeks. Increased level is seen in about 30% of patients with chronic hepatitis C infection . Other
conditions like uncomplicated diabetes mellitus, acute pancreatitis, myocardial infarction,

5. anorexia nervosa, Gullian barre syndrome, hyperthyroidism, obesity and dystrophica myotonica
caused elevated levels of GGT. Elevated serum GGT levels of more than 10 times is observed in
alcoholism. It is partly related to structural liver damage, hepatic microsomal enzyme induction
or alcoholic pancreatic damage. GGT can also be an early marker of oxidative stress since serum
antioxidant carotenoids namely lycopene, α-carotene, β-carotene, and β-cryptoxanthin are
inversely associated with alcohol-induced increase of serum GGT found in moderate and heavy
drinkers . GGT levels may be 2–3 times greater than the upper reference value in more than 50%
of the patients with nonalcoholic fatty liver disease . There is a significant positive correlation
between serum GGT and triglyceride levels in diabetes and the level decreases with treatment
especially when treated with insulin. Whereas serum GGT does not correlate with hepatomegaly
in diabetes mellitus . Serum GGT activity was significantly lower in the second and third
trimesters of normal asymptomatic pregnancy . The levels of GGT in hyperemesis gravidarum
was 45U/L, in pre-eclampsia 17U/L, and 35U/L in hemolysis with low platelet count and
elevated liver enzymes was found during symptomatic pregnancy . The primary usefulness of
GGT is limited in ruling out bone disease as GGT is not found in bone
Requirement
Equipment • Spectrophotometer • Stopwatch or timer
Reagents :
• AST testing kit • ALT testing kit • Sodium hydroxide (0.4 mol/L, 16 g in 1 L distilled water) •
Calibrator • Low and high controls
Procedure:
i.set spectrophotometer at a wavelength of 340nm.
ii.use 1cm by 1cm cuvette.
iii.work at a room temperature
iv.seal the spectrophotometer with distilled water or air.
v.Enzyme is from the plasma given.
vi.Pipette 1ml of working reagent.
vii.add 0.1ml of the sample.
viii.For AST , mix and transfer into the cuvette for measurement after 1minute of incubation, set
a stopwatch at the same time, then take the absorbance.
ix.while the cuvette containing the sample in the spectrophotometer, take measurementof O.D
every 1minute for the next 3minutes.

6. x.Calculate the difference between o.d and average o.d differences per minute.
Xi. To get absorbance per minute take the 1st 2 readings and get their difference.
xii.Repeat the same procedure using ALT working reagent for ALT and compare with their
normal values.
Calculations:
Mean reading= A1 + A2 + A3/3= change of absorbance per minute x factor(1750) to get results
in units/liter(Un/l)
(a) AST: ((1st minute o.d – 2nd minute od)+(2nd minute o.d – 3rd minute od)+(3rd minute o.d -
4thminute o.d))/ 3 x a costant
=(0.005+0.004+0.005)/3 x 1750
=8.1667 units/L
(b)ALT:
=(0.007+0.002+0.004)/3v x 1750
=7.5833units/L
Discusion and Conclusion:
The values obtained for both ALT and AST obtained above fall in the standard normal values.
Laboratory liver tests help to elucidate the alteration of markers which reflect the liver disease.
The assessment of enzyme abnormalities like, the predominant pattern of enzyme alteration, the
magnitude of enzyme alteration in the case of aminotransferases, isolated elevation or in
conjugation with some other parameter, the rate of change and the nature of the course of
alteration or follow up of 6 months to 1–2 years helps in the diagnosis of the disease. But a single
laboratory liver test is of little value in screening for liver disease as many serious liver diseases
may be associated with normal levels and abnormal levels might be found in asymptomatic
healthy individuals. The pattern of enzyme abnormality, interpreted in the context of the
patient’s symptoms can aid in directing the subsequent diagnosis.
However different diseases of the liver will cause different types of damage and will affect liver
function tests accordingly. It is possible to suggest which disease may be present from a liver
function test but these tests are not the conclusive way of diagnosing liver disease. They are
helpful, but only part of the picture. They are also useful for monitoring someone with liver
disease, but are not always accurate.
It is well accepted that in certain liver diseases (hepatitis C, for example) the liver function test
may not accurately show the extent of inflammation or fibrosis (formation of scar tissue),
although this does not apply to the majority of people who have liver disease.

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